Trauma is the leading cause of death in this country in people under the age of 54 years. Except for major head injury, hemorrhage and its consequences are the most important causes of morbidity and mortality in these young, otherwise healthy individuals. This Center Grant proposal seeks to advance our understanding of the molecular mechanisms leading to organ injury and dysfunction following trauma and hemorrhage. Our focus is on the events leading to the activation and propagation of pro-inflammatory and stress signaling pathways that contribute to organ damage and dysfunction post-resuscitation. With this focus we seek to better understand the mechanisms involved in the early organ dysfunction in severely injured patients. It has also become clear that the magnitude of the early inflammatory changes and organ dysfunction contribute to the susceptibility of the trauma victim to delayed infection and sepsis through the activation of counter-inflammatory mediators, rendering the host immunosuppressed. Thus, understanding the inciting events could lead to effective approaches to limit not only the early organ damage but also the susceptibility to late sepsis and multiple organ failure (MOF). We propose that one of the keys to understanding organ injury resulting from hemorrhagic shock is to characterize the earliest molecular events leading to the initiation and propagation of inflammatory changes following the traumatic event. We now know that initiation of inflammatory and stress signaling occurs within minutes of the induction of shock and that the amplification of these responses involves a series of overlapping events. Some of these responses originate within the cells (e.g. production ofredox reactants) while others occur in response to extracellular signals (e.g. circulating hormones or alarm proteins). Because there is a clear temporal sequence to the signaling events, we have found it conceptually useful to separate the hemorrhagic shock insult with or without added tissue trauma into discrete phases; the shock or pre-resuscitation phase, the resuscitation phase, and the late post-resuscitation phase. A premise of this proposal is that molecular events that occur during the shock phase, including upregulation of new genes, will control the post-resuscitation inflammatory response. Another major aspect of our overall hypothesis is that the duration and severity of the shock phase determine the degree of phenotypic changes and hence the intensity of the inflammatory response following resuscitation. We have acquired considerable evidence in support of these original hypotheses. For example, redox events during shock activate MAPK followed by upregulation of several genes (e.g. iNOS, Egr-1). These, in turn, regulate post-resuscitation cytokine production and organ damage. This paradigm, although clearly supported by our data, is too simple. We now have evidence that shock leads to release of stress signals (e.g. RAGE ligands, hyluronic acid) that further amplify the inflammatory response. The magnitude and character of these early events control the balance between predominantly pro-inflammatory (e.g. IL-6, TNF) vs. anti-inflammatory (e.g. IL-10) forces. Both are activated simultaneously but overproduction of IL-6 leads directly to liver and gut damage, while high levels of IL-10 are associated with less inflammation and organ injury. Although the events are time-driven and sometimes simultaneous, it is evident that they are not linear and comprise a response that is characteristic of all complex systems. Although highly integrated and interconnected, there is the potential for considerable redundancy and variability. Our mechanism-driven, reductionist approach has proven useful to identify some of the key pathways and mediators. However, as the quantity of information has expanded, we have recognized the need to integrate this information in a mathematical model that predicts the relationship between key events. Constant refinement of the model will come from the incorporation of new experimental data and validation of experimental data in the clinical setting. Ultimately the model guides the generation of new hypothesis. Just as the early molecular signaling events in trauma and shock are highly integrated so is our research approach. Each of the five projects pursues a defined aspect of the host response to trauma and shock. All of the projects are integrated through well-established collaborative channels and co-reliance on three well-organized cores. Project I (Billiar) pursues the mechanisms involved in the activation of inflammatory pathways in the liver emphasizing the earliest events following the induction of shock and/or trauma. The role of oxygen and nitrogen radicals are emphasized as is the role of rapid response genes such as Egr- 1. Project If (Fink) explores the mechanisms of epithelial cell dysfunction with an emphasis on downstream mediators such as NO, RAGE ligands (alarm proteins) and IL-6. Project III (Bauer) studies the mediators leading to gut motor function failure. Roles for NO and IL-6 have been identified. Recent findings suggest that another alarm molecule released by damaged tissue, hyluronic acid, contribute to the failure through interaction with CD44. Project IV (Pitt) examines the mechanisms leading to early endothelial dysfunction and lung injury. A novel mechanism involving the transcytosis myloperoxidase and of NO-linked albumin is pursued. Project V (Vodovotz/Harbrecht) integrates the experimental findings into a mathematical model of the injury response. Project V also links projects I-VI with the clinical side by gathering clinical data on trauma patients for incorporation into our mathematical model. These collaborations and the overall goal of the center will be promoted significantly by the common use of three well-organized cores. The Animal Models Core (Core B) will provide a source of tissues from animals (both rats and mice) subjected to standardized protocols of hemorrhagic shock, trauma and hemorrhage under the supervision of technically experienced core personnel. This approach maintains consistency of the models between the projects and permits a detailed comparison of results. The Structural Imaging Core (Core C) will provide extraordinary expertise in state-of-the-art tissue imaging, including immunohistochemistry, confocal microscopy, electron microscopy, quantitative morphology, and in situ hybridization. Since each of the projects tests an individual hypothesis that seeks to identify the molecular events in tissues in hemorrhagic or traumatic shock, efficient and accurate structural imaging to localize these changes is essential to each investigator. The Administrative Core (Core A) will provide the critical organization needed to assure productive collaboration and communication. Based on our progress thus far, we are fully confident that our approach will continue to lead to productive collaboration and effective testing of a novel and important hypotheses.